The most effective approach for managing HIV/AIDS is through therapeutic intervention. Current anti-viral drugs target reverse transcription, virus maturation, and entry. Novel drugs against as of yet unexploited targets are essential and any replication step that the virus must accomplish to grow in human cells defines a viable target. After entering a susceptible target cell, the virus must move from the cell periphery, through the cytoplasm, and into the nucleus where it will recombine its neo- synthesized DNA with a cell chromosome. Mutations that impede preintegration trafficking are lethal to the virus, yet the mechanistic details of how the virus moves through the cytoplasm and enters the nucleus are poorly understood. This proposal will uncover essential mechanistic details of HIV-1 preintegration trafficking and nuclear import using a tour de force of molecular techniques. Proteomic approaches will be utilized to uncover essential virus-host interactions. Biochemical fractionation will be utilized to compare the composition of wild-type and mutant replication complexes, and fluorescent confocal microscopy will be used to visualize the complexes as they move through the cell. Fluorescent in situ hybridization will be used to visualize viral nucleic acids once they are inside the nucleus. The results of these experiments will reveal salient features of how the virus moves through the cell to accomplish the essential preintegration tasks in its lifecycle. This knowledge in the long run will define novel targets for therapeutic interaction to develop new anti-viral drugs in the fight against HIV/AIDS.